The overall objective of this work is to investigate the molecular mechanisms that regulate developmental communication between neural and vascular cells as the brain forms. The goal of the proposed research is to investigate the role of vascular endothelial growth factor A (VEGF) and its mechanism of action in regulating early cortical neurogenesis. The results from this study will clarify our understanding of the dual nature of VEGF in its ability to regulate survival and differentiation of vascular and neural cells during neurogenesis. The key to understanding the potential of the nervous system to respond to damage or stress, whether during development or in the adult, lies in the regulatory mechanisms that control neurogenesis in the context of the vasculature. Historically, VEGF has been investigated as a potent regulator of angiogenesis (reviewed in1). However, more recent investigations have suggested an expanded role for VEGF in the nervous system (reviewed in 2, 3). VEGF undergoes alternative splicing to generate isoforms that differ in their size and ability to bind to heparan sulfate proteoglycans in the extracellular matrix and on the cell surface4. This latter property of the isoforms allows VEGF to establish a gradient of diffusible and locally retained signals in the cellular microenvironment. Although a number of studies have suggested a role for VEGF in the nervous system, for example in blood vessel patterning2, 3, little is known about the direct role that VEGF plays in neurogenesis. With previous support from NINDS (1 R15 NS057807-01), we have determined that the three major VEGF isoforms and VEGF receptors are differentially expressed during a key window of neuralepithelial development in the mouse brain. Moreover, a Pax6-positive population of neural stem cells is reduced in a transgenic mouse model expressing only the diffusible VEGF120 isoform or only the locally-retained VEGF188 isoform. As a consequence, the normal expression pattern of Tbr2-positive intermediate progenitor cells, critical for early cortical layer establishment, is disrupted. The proposed research will test the hypothesis that the VEGF isoforms establish a gradient that regulates the proliferation and differentiation of Pax6-positive neural stem cells contributing to normal layer formation in the cortex. The hypothesis will be tested with the following aims. Aim #1. To assess the role of the VEGF isoforms in regulating early cortical development using mice expressing single VEGF isoforms or combinations of VEGF isoforms. Aim #2. To identify the mechanism of VEGF isoform action in a heterotypic cell culture model incorporating neural and vascular cell components. The results of this proposal will be important for identifying how heterotypic cell-cell interactions between the neural and vascular systems regulate neurogenesis in early brain formation. PUBLIC HEALTH RELEVANCE: Neurogenesis and angiogenesis are critical processes in development, but also play key roles in response to injury and pathological stressors, such as cerebral hemorrhage and stroke. It is important to understand not only how neurogenesis and angiogenesis are regulated in concert, but also to determine what the molecular regulators are in this process. As active neural stem cell populations are identified in the adult the possibilities for more novel therapeutic approaches and interventions become available and understanding molecular regulation of neural stem fate will be critical to success. The current proposal will address specifically a role for VEGF in regulating the survival, proliferation, and differentiation of neural stem cells in the context of the developing vasculature.